The objective of this project is to study the regulation of glutamate receptor complexes after a brief period of ischemia followed by reperfusion (I/R). Specifically, we will investigate the protein composition, trafficking, clustering, and signal transduction of NMDA and AMPA receptor complexes associated with ischemia. NMDA and AMPA receptors form clusters or complexes with numerous proteins such as PSD-95 and SynGAP in postsynaptic densities (PSDs). Constituents of these receptor clusters are dynamically regulated during long-term potentiation. However, overactivation of these receptors after I/R leads to excitotoxicity. Knowledge of aberrant glutamate receptor complex organization that leads to excitotoxicity may provide new therapeutic avenues for stroke patients. We have recently established a series of state-of-the-art techniques to study synaptic morphological and molecular modifications, and found dramatic alterations in synaptic ultrastructure, molecular composition, and signal transduction after I/R. Specifically, our studies have clearly demonstrated that recruitment of signaling protein kinases, and accumulation of tyrosine phosphorylated proteins in PSDs, are two of the most dominant molecular events altered in synapses after I/R. Because the effects of I/R are most pronounced on glutamatergic pathways, we will study further the regulation of two key synaptic regulators, the NMDA and AMPA receptor complexes, after I/R in vivo and oxygen glucose deprivation (OGD) in neuronal culture. Aim 1 will test the hypothesis that brain ischemia leads to aberrant reorganization of NMDA and AMPA receptor complexes by means of tyrosine phosphorylation of key synaptic proteins, which contributes to excitotocixity after ischemia. Aim 2 will investigate whether tyrosine phosphorylation of NR2B, PSD95 and SynGAP after an episode of OGD alters their synaptic trafficking and clustering, leading to modification of synaptic morphology and signal transduction in neuronal cultures. These studies will produce comprehensive data for understanding aberrant glutamate receptor organization-mediated excitotoxicity, and thus may provide new therapeutic avenues for stroke patients.